In the medical and biological fields, flow cytometers are widely used. A flow cytometer analyzes the type, frequency, and characteristics of a measurement object such as cells or genes by allowing a photoelectric converter such as a photomultiplier or an avalanche photodiode to receive fluorescence emitted by the measurement object irradiated with laser light.
More specifically, in a flow cytometer, a suspension liquid containing a measurement object obtained by labeling an analyte such as a biological material (e.g., cells, DNA, RNA, enzymes, or proteins) with a fluorescent reagent is allowed to flow through a tube together with a sheath liquid flowing under pressure at a speed of about 10 m/s or less so that a laminar sheath flow is formed. The flow cytometer receives fluorescence emitted by a fluorochrome attached to the measurement object by irradiating the measurement object in the laminar sheath flow with laser light and identifies the analyte by using the fluorescence as a label.
Such a flow cytometer can measure the relative amounts of, for example, DNA, RNA, enzymes, proteins etc. contained in a cell, and also can quickly analyze their properties. Further, a cell sorter or the like is used to identify a specific type of cell or chromosome based on fluorescence and selectively and quickly collect only the identified specific cells or chromosomes alive.
When using such a cell sorter, the flow cytometer is required to quickly identify more kinds of measurement objects with high accuracy based on information about fluorescence.
Patent Document 1 discloses a fluorescence detecting device and a fluorescence detecting method which are capable of quickly identifying many kinds of measurement objects with high accuracy by calculating the fluorescence life (fluorescence relaxation time) of fluorescence emitted by a measurement object irradiated with laser light.
Patent Document 1 describes that the phase delay of a fluorescent signal of fluorescence emitted by a measurement object irradiated with intensity-modulated laser light with respect to a modulation signal used to modulate the intensity of the laser light is determined and the fluorescence relaxation time of the fluorescence is calculated from the phase delay.
However, the device and the method disclosed in Patent Document 1 have the following problem in effectively identifying two or more types of fluorescence emitted by measurement objects.
In order to measure measurement objects that emit two or more types of fluorescence, filters or the like corresponding to the wavelengths of respective fluorescence are used so that different photoelectric converters correspond to different wavelength bands. However, each fluorescence emitted by excitation with laser light has a broad wavelength spectrum, and is therefore received by two or more of the photoelectric converters. That is, a fluorescent signal generated by one photoelectric converter is not always a signal derived just from single fluorescence and may be a signal derived from a mixture of two or more types of fluorescence.
Therefore, when a phase delay is determined using such a fluorescent signal derived from a mixture of two or more types of fluorescence, an accurate fluorescence relaxation time cannot be obtained from such a phase delay.